Project Summary/Abstract Autoimmune diseases appear to be caused by dangerous non-tolerant CD4+ T cells specific for an affected individual's own proteins. Current therapies to control these aberrant T cell responses are unsatisfactory because they inhibit all T cells, including helpful ones specific for pathogens and cancer cells. The question now before the field is how to induce self-peptide specific immunological tolerance in CD4+ T cells that cause autoimmune disease. Anergy induction in dangerous CD4 T cells and peripheral regulatory T (Treg) cell expansion represent two potentially important therapeutic strategies to durably control autoimmune disease. Research during the last grant period established that anergy naturally occurs in the polyclonal CD4 repertoire of healthy mice in association with the up-regulation of CD73, FR4, and Nrp1. Remarkably, anergic conventional Foxp3? CD44hi Nrp1+ FR4+ CD73+ CD4 T cells were also observed to differentiate into functional Foxp3+ Treg cells when adoptively transferred into Treg-deficient Tcra?/? hosts. Therefore, this discovery of `anergy-derived' Treg cells offers a new avenue for the design of Treg cell therapies to control CD4 T cell-mediated autoimmunity in an antigen-specific fashion. Research proposed in this application intends to characterize many of the factors that govern natural anergy induction and the generation of Treg progenitors, including autoreactive TCRs, tissue-restricted target self- antigens, tolerogenic signaling events, CpG DNA de-methylations, and anergic gene expression signatures. Furthermore, proposed experiments will determine how anergy-derived Foxp3+ Treg cells can be generated in normal hosts and investigate their capacity to suppress autoimmune disease. Finally, experiments will establish the feasibility of investigating the immunology of anergy and anergy-derived Treg cells in humans.